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Influence of extenders and cooling rates on epididymal sperm of Lewis rat strain

Yıl 2015, , 57 - 62, 01.03.2015

Ömer Varışlı Cansu Agca Yüksel Agca

https://doi.org/10.1501/Vetfak_0000002658

Öz

The objective of this study was to determine the appropriate cooling rates and extender for improving the freezability of epididymal sperm of Lewis rat strain. The epididymal sperm from Lewis rats were suspended in different freezing extenders which were HEPES buffered Tyrode’s lactate (TL-HEPES), modified Kreb’s Ringer bicarbonate (mKRB), 3% dehydrated skim milk (SM), Salamon’s Tris-citrate (TRIS), and TES/Tris (TES). The extenders contained 20% (v/v) egg yolk (EY), 0.75% equex paste (EP) and 0.1 M raffinose or 0.1 M sucrose. The sperm samples were frozen by automated freezer, using various cooling rates (10, 40, 70, and 100 °C/min). After thawing, sperm samples were examined for integrity of plasma and acrosomal integrity, and mitochondrial membrane potential (MMP). Type of extenders had significant effect on cryosurvival (p<0.05). The post-thaw highest motility (27.5%) was achieved for the sperm samples frozen in the presence of TES extender (p<0.05). As a result, although cooling rate did not significantly affect cryopreservation, the extenders affected post-thaw sperm parameters and TES considerably improved the freezability of epididymal Lewis rat sperm

Anahtar Kelimeler

Cooling rate epididymal sperm extenders freezing Lewis rat strain

Kaynakça

  • Agca Y, Critser JK (2005): Assisted reproductive technologies and genetic engineering in rats. 165-190. In: MA Suckow, SH Weisbroth, CL Franklin (Eds), The Laboratory Rat. Second edition, San Diego (CA).
  • Aisen E, Quintana M, Medina V, Morello H, Venturino A (2005): Ultramicroscopic and biochemical changes in ram spermatozoa cryopreserved with trehalose-based hypertonic extenders. Cryobiology, 50, 239–249.
  • Arriola J, Foote RH (1987): Glycerolation and thawing effects on bull spermatozoa frozen in detergent-treated egg yolk and whole egg extenders. J Dairy Sci, 70, 1664–1670.
  • Axnér E, Hermansson U, Linde-Forsberg C (2004): The effect of Equex STM paste and sperm morphology on post- thaw survival of cat epididymal spermatozoa. Anim Reprod Sci, 84, 179–191.
  • Barbas JP, Mascarenhas RD (2009): Cryopreservation of domestic animal sperm cells. Cell Tissue Bank, 10, 49– 62.
  • Batellier F, Vidament M, Fauquant J, Duchamp G, Arnaud G, Yvon JM, Magistrini M (2001): Advances in cooled semen technology. Anim Reprod Sci, 68, 181–190.
  • Bavister BD, Leibfred ML, Lieberman G (1983): Development of preimplantation embryos of the golden hamster in a defined medium. Biol Reprod, 28, 235–247.
  • Carvajal G, Cuello C, Ruiz M, Vazquez JM, Martinez EA, Roca J (2004): Effects of centrifugation before freezing on boar sperm cryosurvival. J Androl, 25, 389–396.
  • Curry MR (2007): Cryopreservation of mammalian semen. Methods Mol Biol, 368, 303-311.
  • Curry MR, Millar JD, Watson PF (1994): Calculated optimal cooling rates for ram and human sperm, cryopreservation fail to conform with empirical observations. Biol Reprod, 51, 1014–1021.
  • Devireddy RV, Swanlund DJ, Roberts KP, Bischof JC (1999): Subzero water permeability parameters of mouse spermatozoa in the presence of extracellular ice and cryoprotective agents. Biol Reprod, 61, 764–775.
  • Fiser PS, Fairfull RW (1990): Combined effect of glycerol concentration and cooling velocity on motility and acrosome integrity of boar spermatozoa frozen in 0.5 ml straws. Mol Reprod Dev, 25, 123-129.
  • Gao D, Mazur P, Critser JK (1997): Fundamental cryobiology of mammalian spermatozoa. 263-328. In: AM Karow, JK Critser (Eds), Reproductive Tissue Banking Scientific Principles. Academic Press, San Diego (CA).
  • Hagiwara M, Choi JH, Devireddy RV, Roberts KP, Wolkers WF, Makhlouf A (2009): Cellular biophysics during freezing of rat and mouse sperm predicts post-thaw motility. Biol Reprod, 81, 700–706.
  • Hammerstedt RH, Graham JK, Nolan JP (1990): Cryopreservation of mammalian sperm: what we ask them to survive. J Androl, 11, 73–88.
  • Henry MA, Noiles EE, Gao D, Mazur P, Critser JK (1993): Cryopreservation of human spermatozoa. IV. The effects of cooling rate and warming rate on the maintenance of motility, plasma membrane integrity, and mitochondrial function. Fertil Steril, 60, 911-918.
  • Hirabayash M, Kato M, Aoto T, Sekimoto A, Ueda M, Miyoshi I, Kasai N, Hochi S (2002): Offspring derived from intracytoplasmic injection of transgenic rat sperm. Transgenic Res, 11, 221-228.
  • Hochi S, Watanabe K, Kato M, Hirabayashi M (2008): Live rats resulting from injection of oocytes with spermatozoa freeze-dried and stored for one year. Mol Reprod Dev, 75, 890-894.
  • Holt WV (2000): Fundamental aspects of sperm cryobiology: the importance of species and individual differences. Theriogenology, 53, 47–58.
  • National Research Council (1996): Animal environment, housing, and management. 21-55. In: Guide for the Care and Use of Laboratory Animals. National Academy, Press Washington (DC).
  • Kasai T, Ogawa K, Mizuno K, Nagai S, Uchida Y, Ohta S, Fujie M, Suzuki K, Hirata S, Hoshi K (2002): Relationship between sperm mitochondrial membrane potential, sperm motility, and fertility potential. Asian J Androl, 4, 97-103.
  • Katkov II, Mazur P (1999): Factors affecting yield and survival of cells when suspensions are subjected to centrifugation: I. Influence of acceleration, time of centrifugation, and length of the suspension column in homogeneous centrifugal fields. Cell Biochem Biophys, 31, 231–245.
  • Kim S, Agca C, Agca Y (2012): Changes in rat spermatozoa function after cooling, cryopreservation and centrifugation processes. Cryobiology 65, 215-23.
  • Koshimoto C, Gamliel E, Mazur P (2000): Effect of osmolality and oxygen tension on the survival of mouse sperm frozen to various temperatures in various concentrations of glycerol and raffinos. Cryobiology, 41, 204-231.
  • Koshimoto C, Mazur P (2002): Effects of cooling and warming rate to and from -70 degrees C, and effect of further cooling from -70 to -196 degrees C on the motility of mouse spermatozoa. Biol Reprod, 66, 1477-1484.
  • Lazar J, Moreno C, Jacob HJ, Kwitek AE (2005): Impact of genomics on research in the rat. Genome Res, 15, 1717–1728.
  • Mazur P (1970): Cryobiology: the freezing of biological systems. Science, 168, 939–949.
  • Mazur P (1984): Freezing of living cells: mechanisms and implications. Am J Physiol, 247, 125-142.
  • Nakagata N (2000): Cryopreservation of mouse spermatozoa. Mamm Genome, 11, 572–576.
  • Nakagata N, Takeshima T (1992): High fertilizing ability of mouse spermatozoa diluted slowly after cryopreservation. Theriogenology, 37, 1283–1291.
  • Nakatsukasa E, Inomata T, Ikeda T, Shino M, Kashiwazaki N (2001): Generation of live rat offspring by intrauterine insemination with epididymal spermatozoa cryopreserved at -196 °C. Reproduction, 122, 463–467.
  • Nakatsukasa E, Kashiwazaki N, Takizawa A, Shino M, Kitada K, Serikawa T, Hakamata Y, Kobayashi E, Takahashi R, Ueda M, Nakashima T, Nakagata N (2003): Cryopreservation of spermatozoa from closed colonies, and inbred, spontaneous mutant, and transgenic strains of rats. Comp Med, 53, 639–641.
  • Oh SH, Miyoshi K, Funahashi H (1998): Rat oocytes fertilized in modified rat 1 cell embryo culture medium containing a high sodium chloride concentration and bovine serum albumin maintain developmental ability to the blastocyst stage. Biol Reprod, 59, 884–889.
  • Okuyama M, Isogai S, Saga M, Hamada H, Ogawa S (1990): In vitro fertilization (IVF) and artificial insemination by cryopreserved spermatozoa in mouse. J Fertil Implant, 7, 116–119.
  • Parks J (1997): Hypothermia and mammalian gametes. 229-260. In: A Karow, JK Critser (Eds), Reproductive Tissue Banking, Scientific Principles. Academic Press, San Diego (CA).
  • Pena A, Linde-Forsberg C (2000): Effects of equex, one- or two- step dilution, and two freezing and thawing rates on post-thaw survival of dog spermatozoa. Theriogenology, 54, 859–875.
  • Penfold LM, Moore HD (1993): A new method for cryopreservation of mouse spermatozoa. J Reprod Fertil, 99, 131–134.
  • Purdy PH (2006): A review on goat sperm cryopreservation. Small Rum Research, 63, 215–225.
  • Salamon S, Maxwell WM (2000): Storage of ram semen. Anim Reprod Sci, 18, 77-111.
  • Schreuders PD, Jetton AE, Baker JL, Critser JK, Mazur P (1996): Mechanical and chill sensitivity of mouse sperm. Cryobiology, 33, 676–677.
  • Si W, Benson JD, Men H, Critser JK (2006): Osmotic tolerance limits and effects of cryoprotectants on the motility, plasma membrane integrity and acrosomal integrity of rat sperm. Cryobiology, 53, 336–348.
  • Stacy R, Eroglu A, Fowler A, Biggers J, Toner M (2006): Thermal characterization of nakagata's mouse sperm freezing protocol. Cryobiology, 52, 99-107.
  • Tada N, Sato M, Yamanoi J, Mizorogi T, Kasai K, Ogawa S (1990): Cryopreservation of mouse spermatozoa in the presence of raffinose and glycerol. J Reprod Fertil, 89, 511–516.
  • Tesson L, Cozzi J, Menoret S, Remy S, Usal C, Fraichard A, Anegon I (2005): Transgenic modifications of the rat genome. Transgenic Res, 14, 531–546.
  • Toyoda Y, Chang MC (1968): Sperm penetration of rat eggs in vitro after dissolution of zona pellucida by chymotrypsin. Nature, 220, 589–591.
  • Varisli O, Scott H, Agca C, Agca Y (2013): The effects of cooling rates and type of freezing extenders on cryosurvival of rat sperm. Cryobiology, 67, 109-116.
  • Varisli O, Uguz C, Agca C, Agca Y (2009): Effect of chilling on the motility and acrosomal integrity of rat sperm in the presence of various extenders. J Am Assoc Lab Anim, 48, 1-7.
  • Varisli O, Uguz C, Agca C, Agca Y (2009): Various physical stress factors on rat sperm motility, integrity of acrosome, and plasma membrane. J Androl, 30, 75–86.
  • Watson PF (1995): Recent developments and concepts in the cryopreservation of spermatozoa and the assessment of their post-thawing function. Reprod Fertil Dev, 7, 871– 891.
  • Watson PF (2000): The causes of reduced fertility with cryopreserved semen. Anim Reprod Sci, 60-61, 481–492.
  • Woelders H, Matthijs A, Engel B (1997): Effects of trehalose and sucrose, osmolality of the freezing medium and cooling rate on viability and intactness of bull sperm after freezing and thawing. Cryobiology, 35, 93–105.
  • Yamashiro H, Han YJ, Sugawara A, Tomioka I, Hoshino Y, Sato E (2007): Freezability of rat epididymal sperm induced by raffinose in modified Krebs-Ringer bicarbonate (mKRB) based extender solution. Cryobiology, 55, 285–294.
  • Yamashiro H, Kumamoto K, Wang H, Yamashita Y, Terada T (2006): Effect of semen collection in extender solution on the characteristics of goat spermatozoa. J Reprod Dev, 52, 397–406.
  • Yamashiro H, Toyomizu M, Toyama N, Aono N,
  • Sakurai M, Hiradate Y, Yokoo M, Moisyadi S, Sato E (2010): Extracellular ATP and dibutyryl cAMP enhance the freezability of rat epididymal sperm. J Am Assoc Lab Anim Sci, 49, 167–172.

Lewis ırkı rat sperması üzerine değişik sulandırıcı ve dondurma hızının etkisi

Yıl 2015, , 57 - 62, 01.03.2015

Ömer Varışlı Cansu Agca Yüksel Agca

https://doi.org/10.1501/Vetfak_0000002658

Öz

Çalışmanın amacı Lewis ırkı rat sperması için, uygun dondurma hızı ve sulandırıcılar tespit ederek spermanın dondurulabilirliğinin artırılmasıdır. Ratlardan alınan epididimal sperma HEPES buffered Tyrode’s lactate (TL-HEPES), modified Kreb’s Ringer bicarbonate (mKRB), %3 süt tozu (SM), Tris-citrate (TRIS) ve Tris/TES (TES) solüsyonları ile sulandırıldı. Sulandırıcılara 20% (v/v) yumurta sarısı, 0.75% Equex Paste ve 0.1 M rafinoz yada 0.1 M sükroz ilave edildi. Sperma örnekleri kontrollü dondurucu kullanılarak değişik soğutma hızında (10, 40, 70 ve 100 °C/dk) donduruldu. Dondurma sonrası spermalar plazma, akrozom ve mitokondri membran bütünlüğü yönünden değerlendirildi. Sulandırıcı tipinin dondurmada etkili olduğu saptandı (p<0.05). Dondurma-çözüm işlemi sonrası en yüksek motilite (%27.5) TES sulandırıcısından elde edildi (p<0.05). Sonuç olarak, dondurma hızının sperm dondurulmasına önemli bir etkisi tespit edilemese de, sulandırıcıların dondurma-çözüm sonrası sperm parametrelerini etkilediği ve TES’in epididimal Lewis rat sperması dondurulabilirliğini, önemli derecede geliştirdiği saptandı

Anahtar Kelimeler

Dondurma dondurma hızı epididimal sperma sulandırıcı Lewis rat ırkı

Kaynakça

  • Agca Y, Critser JK (2005): Assisted reproductive technologies and genetic engineering in rats. 165-190. In: MA Suckow, SH Weisbroth, CL Franklin (Eds), The Laboratory Rat. Second edition, San Diego (CA).
  • Aisen E, Quintana M, Medina V, Morello H, Venturino A (2005): Ultramicroscopic and biochemical changes in ram spermatozoa cryopreserved with trehalose-based hypertonic extenders. Cryobiology, 50, 239–249.
  • Arriola J, Foote RH (1987): Glycerolation and thawing effects on bull spermatozoa frozen in detergent-treated egg yolk and whole egg extenders. J Dairy Sci, 70, 1664–1670.
  • Axnér E, Hermansson U, Linde-Forsberg C (2004): The effect of Equex STM paste and sperm morphology on post- thaw survival of cat epididymal spermatozoa. Anim Reprod Sci, 84, 179–191.
  • Barbas JP, Mascarenhas RD (2009): Cryopreservation of domestic animal sperm cells. Cell Tissue Bank, 10, 49– 62.
  • Batellier F, Vidament M, Fauquant J, Duchamp G, Arnaud G, Yvon JM, Magistrini M (2001): Advances in cooled semen technology. Anim Reprod Sci, 68, 181–190.
  • Bavister BD, Leibfred ML, Lieberman G (1983): Development of preimplantation embryos of the golden hamster in a defined medium. Biol Reprod, 28, 235–247.
  • Carvajal G, Cuello C, Ruiz M, Vazquez JM, Martinez EA, Roca J (2004): Effects of centrifugation before freezing on boar sperm cryosurvival. J Androl, 25, 389–396.
  • Curry MR (2007): Cryopreservation of mammalian semen. Methods Mol Biol, 368, 303-311.
  • Curry MR, Millar JD, Watson PF (1994): Calculated optimal cooling rates for ram and human sperm, cryopreservation fail to conform with empirical observations. Biol Reprod, 51, 1014–1021.
  • Devireddy RV, Swanlund DJ, Roberts KP, Bischof JC (1999): Subzero water permeability parameters of mouse spermatozoa in the presence of extracellular ice and cryoprotective agents. Biol Reprod, 61, 764–775.
  • Fiser PS, Fairfull RW (1990): Combined effect of glycerol concentration and cooling velocity on motility and acrosome integrity of boar spermatozoa frozen in 0.5 ml straws. Mol Reprod Dev, 25, 123-129.
  • Gao D, Mazur P, Critser JK (1997): Fundamental cryobiology of mammalian spermatozoa. 263-328. In: AM Karow, JK Critser (Eds), Reproductive Tissue Banking Scientific Principles. Academic Press, San Diego (CA).
  • Hagiwara M, Choi JH, Devireddy RV, Roberts KP, Wolkers WF, Makhlouf A (2009): Cellular biophysics during freezing of rat and mouse sperm predicts post-thaw motility. Biol Reprod, 81, 700–706.
  • Hammerstedt RH, Graham JK, Nolan JP (1990): Cryopreservation of mammalian sperm: what we ask them to survive. J Androl, 11, 73–88.
  • Henry MA, Noiles EE, Gao D, Mazur P, Critser JK (1993): Cryopreservation of human spermatozoa. IV. The effects of cooling rate and warming rate on the maintenance of motility, plasma membrane integrity, and mitochondrial function. Fertil Steril, 60, 911-918.
  • Hirabayash M, Kato M, Aoto T, Sekimoto A, Ueda M, Miyoshi I, Kasai N, Hochi S (2002): Offspring derived from intracytoplasmic injection of transgenic rat sperm. Transgenic Res, 11, 221-228.
  • Hochi S, Watanabe K, Kato M, Hirabayashi M (2008): Live rats resulting from injection of oocytes with spermatozoa freeze-dried and stored for one year. Mol Reprod Dev, 75, 890-894.
  • Holt WV (2000): Fundamental aspects of sperm cryobiology: the importance of species and individual differences. Theriogenology, 53, 47–58.
  • National Research Council (1996): Animal environment, housing, and management. 21-55. In: Guide for the Care and Use of Laboratory Animals. National Academy, Press Washington (DC).
  • Kasai T, Ogawa K, Mizuno K, Nagai S, Uchida Y, Ohta S, Fujie M, Suzuki K, Hirata S, Hoshi K (2002): Relationship between sperm mitochondrial membrane potential, sperm motility, and fertility potential. Asian J Androl, 4, 97-103.
  • Katkov II, Mazur P (1999): Factors affecting yield and survival of cells when suspensions are subjected to centrifugation: I. Influence of acceleration, time of centrifugation, and length of the suspension column in homogeneous centrifugal fields. Cell Biochem Biophys, 31, 231–245.
  • Kim S, Agca C, Agca Y (2012): Changes in rat spermatozoa function after cooling, cryopreservation and centrifugation processes. Cryobiology 65, 215-23.
  • Koshimoto C, Gamliel E, Mazur P (2000): Effect of osmolality and oxygen tension on the survival of mouse sperm frozen to various temperatures in various concentrations of glycerol and raffinos. Cryobiology, 41, 204-231.
  • Koshimoto C, Mazur P (2002): Effects of cooling and warming rate to and from -70 degrees C, and effect of further cooling from -70 to -196 degrees C on the motility of mouse spermatozoa. Biol Reprod, 66, 1477-1484.
  • Lazar J, Moreno C, Jacob HJ, Kwitek AE (2005): Impact of genomics on research in the rat. Genome Res, 15, 1717–1728.
  • Mazur P (1970): Cryobiology: the freezing of biological systems. Science, 168, 939–949.
  • Mazur P (1984): Freezing of living cells: mechanisms and implications. Am J Physiol, 247, 125-142.
  • Nakagata N (2000): Cryopreservation of mouse spermatozoa. Mamm Genome, 11, 572–576.
  • Nakagata N, Takeshima T (1992): High fertilizing ability of mouse spermatozoa diluted slowly after cryopreservation. Theriogenology, 37, 1283–1291.
  • Nakatsukasa E, Inomata T, Ikeda T, Shino M, Kashiwazaki N (2001): Generation of live rat offspring by intrauterine insemination with epididymal spermatozoa cryopreserved at -196 °C. Reproduction, 122, 463–467.
  • Nakatsukasa E, Kashiwazaki N, Takizawa A, Shino M, Kitada K, Serikawa T, Hakamata Y, Kobayashi E, Takahashi R, Ueda M, Nakashima T, Nakagata N (2003): Cryopreservation of spermatozoa from closed colonies, and inbred, spontaneous mutant, and transgenic strains of rats. Comp Med, 53, 639–641.
  • Oh SH, Miyoshi K, Funahashi H (1998): Rat oocytes fertilized in modified rat 1 cell embryo culture medium containing a high sodium chloride concentration and bovine serum albumin maintain developmental ability to the blastocyst stage. Biol Reprod, 59, 884–889.
  • Okuyama M, Isogai S, Saga M, Hamada H, Ogawa S (1990): In vitro fertilization (IVF) and artificial insemination by cryopreserved spermatozoa in mouse. J Fertil Implant, 7, 116–119.
  • Parks J (1997): Hypothermia and mammalian gametes. 229-260. In: A Karow, JK Critser (Eds), Reproductive Tissue Banking, Scientific Principles. Academic Press, San Diego (CA).
  • Pena A, Linde-Forsberg C (2000): Effects of equex, one- or two- step dilution, and two freezing and thawing rates on post-thaw survival of dog spermatozoa. Theriogenology, 54, 859–875.
  • Penfold LM, Moore HD (1993): A new method for cryopreservation of mouse spermatozoa. J Reprod Fertil, 99, 131–134.
  • Purdy PH (2006): A review on goat sperm cryopreservation. Small Rum Research, 63, 215–225.
  • Salamon S, Maxwell WM (2000): Storage of ram semen. Anim Reprod Sci, 18, 77-111.
  • Schreuders PD, Jetton AE, Baker JL, Critser JK, Mazur P (1996): Mechanical and chill sensitivity of mouse sperm. Cryobiology, 33, 676–677.
  • Si W, Benson JD, Men H, Critser JK (2006): Osmotic tolerance limits and effects of cryoprotectants on the motility, plasma membrane integrity and acrosomal integrity of rat sperm. Cryobiology, 53, 336–348.
  • Stacy R, Eroglu A, Fowler A, Biggers J, Toner M (2006): Thermal characterization of nakagata's mouse sperm freezing protocol. Cryobiology, 52, 99-107.
  • Tada N, Sato M, Yamanoi J, Mizorogi T, Kasai K, Ogawa S (1990): Cryopreservation of mouse spermatozoa in the presence of raffinose and glycerol. J Reprod Fertil, 89, 511–516.
  • Tesson L, Cozzi J, Menoret S, Remy S, Usal C, Fraichard A, Anegon I (2005): Transgenic modifications of the rat genome. Transgenic Res, 14, 531–546.
  • Toyoda Y, Chang MC (1968): Sperm penetration of rat eggs in vitro after dissolution of zona pellucida by chymotrypsin. Nature, 220, 589–591.
  • Varisli O, Scott H, Agca C, Agca Y (2013): The effects of cooling rates and type of freezing extenders on cryosurvival of rat sperm. Cryobiology, 67, 109-116.
  • Varisli O, Uguz C, Agca C, Agca Y (2009): Effect of chilling on the motility and acrosomal integrity of rat sperm in the presence of various extenders. J Am Assoc Lab Anim, 48, 1-7.
  • Varisli O, Uguz C, Agca C, Agca Y (2009): Various physical stress factors on rat sperm motility, integrity of acrosome, and plasma membrane. J Androl, 30, 75–86.
  • Watson PF (1995): Recent developments and concepts in the cryopreservation of spermatozoa and the assessment of their post-thawing function. Reprod Fertil Dev, 7, 871– 891.
  • Watson PF (2000): The causes of reduced fertility with cryopreserved semen. Anim Reprod Sci, 60-61, 481–492.
  • Woelders H, Matthijs A, Engel B (1997): Effects of trehalose and sucrose, osmolality of the freezing medium and cooling rate on viability and intactness of bull sperm after freezing and thawing. Cryobiology, 35, 93–105.
  • Yamashiro H, Han YJ, Sugawara A, Tomioka I, Hoshino Y, Sato E (2007): Freezability of rat epididymal sperm induced by raffinose in modified Krebs-Ringer bicarbonate (mKRB) based extender solution. Cryobiology, 55, 285–294.
  • Yamashiro H, Kumamoto K, Wang H, Yamashita Y, Terada T (2006): Effect of semen collection in extender solution on the characteristics of goat spermatozoa. J Reprod Dev, 52, 397–406.
  • Yamashiro H, Toyomizu M, Toyama N, Aono N,
  • Sakurai M, Hiradate Y, Yokoo M, Moisyadi S, Sato E (2010): Extracellular ATP and dibutyryl cAMP enhance the freezability of rat epididymal sperm. J Am Assoc Lab Anim Sci, 49, 167–172.
Toplam 55 adet kaynakça vardır.

Ayrıntılar

Birincil Dil İngilizce
Konular Veteriner Cerrahi
Diğer ID JA86TY96UA
Bölüm Araştırma Makalesi
Yazarlar

Ömer Varışlı

Cansu Agca

Yüksel Agca

Yayımlanma Tarihi 1 Mart 2015
Yayımlandığı Sayı Yıl 2015

Kaynak Göster

APA Varışlı, Ö., Agca, C., & Agca, Y. (2015). Influence of extenders and cooling rates on epididymal sperm of Lewis rat strain. Ankara Üniversitesi Veteriner Fakültesi Dergisi, 62(1), 57-62. https://doi.org/10.1501/Vetfak_0000002658
AMA Varışlı Ö, Agca C, Agca Y. Influence of extenders and cooling rates on epididymal sperm of Lewis rat strain. Ankara Univ Vet Fak Derg. Mart 2015;62(1):57-62. doi:10.1501/Vetfak_0000002658
Chicago Varışlı, Ömer, Cansu Agca, ve Yüksel Agca. “Influence of Extenders and Cooling Rates on Epididymal Sperm of Lewis Rat Strain”. Ankara Üniversitesi Veteriner Fakültesi Dergisi 62, sy. 1 (Mart 2015): 57-62. https://doi.org/10.1501/Vetfak_0000002658.
EndNote Varışlı Ö, Agca C, Agca Y (01 Mart 2015) Influence of extenders and cooling rates on epididymal sperm of Lewis rat strain. Ankara Üniversitesi Veteriner Fakültesi Dergisi 62 1 57–62.
IEEE Ö. Varışlı, C. Agca, ve Y. Agca, “Influence of extenders and cooling rates on epididymal sperm of Lewis rat strain”, Ankara Univ Vet Fak Derg, c. 62, sy. 1, ss. 57–62, 2015, doi: 10.1501/Vetfak_0000002658.
ISNAD Varışlı, Ömer vd. “Influence of Extenders and Cooling Rates on Epididymal Sperm of Lewis Rat Strain”. Ankara Üniversitesi Veteriner Fakültesi Dergisi 62/1 (Mart 2015), 57-62. https://doi.org/10.1501/Vetfak_0000002658.
JAMA Varışlı Ö, Agca C, Agca Y. Influence of extenders and cooling rates on epididymal sperm of Lewis rat strain. Ankara Univ Vet Fak Derg. 2015;62:57–62.
MLA Varışlı, Ömer vd. “Influence of Extenders and Cooling Rates on Epididymal Sperm of Lewis Rat Strain”. Ankara Üniversitesi Veteriner Fakültesi Dergisi, c. 62, sy. 1, 2015, ss. 57-62, doi:10.1501/Vetfak_0000002658.
Vancouver Varışlı Ö, Agca C, Agca Y. Influence of extenders and cooling rates on epididymal sperm of Lewis rat strain. Ankara Univ Vet Fak Derg. 2015;62(1):57-62.